Toxoplasma gondii is an obligate intracellular parasite that is responsible for a wide spectrum of disease affecting humans and many animals. In immunoincompetent host, disease could be very severe. T. gondii is now a major cause of opportunistic infection in people with AIDS. There is an urgent need for alternative drugs for the treatment of human toxoplasmosis. Studies of the basic biochemistry of T. gondii could lead to the rational design of new and more effective drugs. We have identified an acidic organelle rich in calcium possibly involved in the regulation of Ca2+ and pH homeostasis in T. gondii that we named the acidocalcisome. We also found that a pyrophosphate is apparently associated with these organelles. Inorganic pyrophosphate occurs in these parasites at concentrations higher than ATP. In addition, we found a "plant-like" enzymatic activity possibly associated with the same organelle in T. gondii: a H+-translocating pyrophosphatase. These results could have important implications because the novel metabolic pathways identified in Toxoplasma could be targets for chemotherapeutic intervention. We intend to focus on studying the acidocalcisomes with the following specific aims: specific aim 1: identification and characterization of the pyrophosphate present in acidocalcisomes; specific aim 2: characterization of T. gondii H+-pyrophosphatase; specific aim 3: investigation of the role of acidocalcisomes in intracellular Ca2+ and pH homeostasis.